Menkes' and Wilson's disease: Study of copper in skin fibroblasts

Abstract

MATERIALS AND METHODSSkin fibroblasts were cultivated in Ham F10 medium supplemented with 15 % fetal calf serum, antibioticantimycotic solution, in 5% carbon dioxide at 37°C. Cells were harvested by trypsin, rinsed three times, suspended in deionized water and lysed by freezing and thawing in an acetone dry-ice bath. Lysates were centrifuged at 2000 g and supernatants were collected for further investigation. Protein concentration was determined by the method of Lowry and copper and zinc by flameless atomic absorption.